Abstract
Purpose :
The lacrimal gland is a tear-secreting organ that prevents dryness of the eyes. Seminal work established that fibroblast growth factor (FGF) signaling is essential for lacrimal gland development. Phospholipase C-γ (PLC-γ) is one of the handful substrates activated by FGF receptors through direct binding, but the physiological function of PLC-γ in lacrimal gland development is largely unexplored.
Methods :
We generated conditional knockout mouse lines using Le-Cre, which is specifically expressed in the lacrimal gland epithelium and linked to an IRES-GFP reporter. We generated systemic mutant mouse lines by CRISPR/Cas9 genome editing techniques. GFP fluorescence and carmine staining were used to visualize lacrimal gland structure.
Results :
Conditional knockout of PLC-γ1 caused lacrimal gland hyperplasia with ectopic branches from the primary duct as well as extra budding from the ventral side of the eye, consistent with a gain-of-function phenotype of FGF signaling. Immunostaining revealed an early onset of lacrimal gland differentiation in both acini and ducts. To investigate the mechanism by which PLC-γ is activated by FGFR signaling in vivo, we generated Fgfr2Y788F allele that lacks the PLC-γ binding site. Mouse carrying homozygous Fgfr2Y788F recapitulated the phenotype of extra-branching associated with PLC-γ1 mutant. In addition, inactivation of the PLC-γ N-terminal SH2 domain (PLC-γR586A) or the C-terminal SH2 domain (PLC-γR694A) mimicked PLC-γ null phenotype, suggesting that both SH2 domains are critical for PLC-γ function in lacrimal gland development.
Conclusions :
Our study suggests that PLC-γ mediates FGFR signaling through both SH2 domains in lacrimal gland development.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.